FEC475.fm Page 758 Thursday, November 16, 2000 8:51 AM
Functional
Ecology 2000
14, 758 –765
TECHNICAL REPORT
Oxford,
Functional
FEC
Blackwell
620269-8463
14
Physiological
475
B.
000
Blackwell
A. Vanderkist
UK
Science,
Ecology
Science,
assessment
etLtd
al.
Ltd of reproductive state and breeding chronology
Graphicraft Limited, Hong Kong
000
Indirect, physiological assessment of reproductive state
and breeding chronology in free-living birds: an example
in the Marbled Murrelet (Brachyramphus marmoratus)
B. A. VANDERKIST,* T. D. WILLIAMS,† D. F. BERTRAM,†‡
L. W. LOUGHEED† and J. L. RYDER†
*Department of Molecular Biology and Biochemistry, 8888 University Drive, Simon Fraser University, Burnaby,
British Columbia, Canada V5A 1S6, †Department of Biological Sciences, 8888 University Drive, Simon Fraser
University, Burnaby, British Columbia, Canada V5A 1S6, and ‡Canadian Wildlife Service, Pacific and Yukon
Region, 5421 Robertson Road, RR # 1, Delta, British Columbia, Canada V4K 3N2
Summary
1. An indirect, physiological method to assess reproductive state in individuals of
unknown status is described. The plasma levels of two main yolk precursors, vitellogenin (VTG) and very-low density lipoproteins (VLDL), are focused on as indices
of egg production, for the characterization of fecund females.
2. Data for a species where breeding chronology could be directly assessed, at the
population level (Cassin’s Auklet, Ptychoramphus aleuticus), confirmed the validity of
this approach: plasma VTG levels were highest during the defined egg-laying period,
and the highest proportion of females were defined as ‘egg-producing’ in this period.
3. Analysis of samples for Marbled Murrelets (Brachyramphus marmoratus) caught
off-nest (i.e. where all individuals were of unknown status), clearly identified a putative
egg-laying phase, with a single, protracted laying period (cf. multiple-broodiness).
4. Analysis of body mass confirmed our characterization of ‘egg-producing’ females:
birds with elevated plasma VTG were on average 40 g heavier than other females,
equivalent to the mass of the single egg (36–41 g).
5. Indirect, physiological assessment of reproductive state provided valuable information on the breeding biology of Marbled Murrelets which would have been difficult to
obtain in any other way (e.g. proportion of fecund females, breeding phenology, single
vs multiple-clutch breeding pattern). Despite some limitations, this technique should
be applicable to any oviparous vertebrate population where essential information
on breeding biology cannot be obtained by more traditional methods.
Key-words: Physiological methods, Ptychoramphus, vitellogenin
Functional Ecology (2000) 14, 758 –765
Introduction
© 2000 British
Ecological Society
Many life-history traits and behavioural decisions are
state-dependent (e.g. McNamara & Houston 1992;
Houston 1993), and consequently it is important to
know the state of individuals in any particular study
(this might be their physiological state or ‘condition’,
migratory state or reproductive state). In the case of
reproductive state or breeding stage this process is
relatively straightforward for most species: investigators can simply locate large numbers of breeding individuals and identify breeding state by direct
observation (see Clutton-Brock, Guiness & Albon
1982; Cooke, Rockwell & Lank 1995, for excellent
examples). This also allows direct assessment of other
key reproductive parameters such as breeding phenology
(onset and duration of the reproductive phase)
and, potentially, population structure (sex ratio, nonbreeders vs breeders, or proportion of fecund females).
Some species cannot be studied in this way, however,
either because they are too cryptic, because they use
remote breeding habitats, or because disturbance at
breeding sites must be minimized (as in the case of
rare or endangered species).
Recently, there has been much interest in the use of
physiological methods to assess physiological state
(namely ‘condition’, reviewed in Brown 1996; immune
function, Ots, Muramagi & Horak 1998), or migratory
state (Jenni-Eiermann & Jenni 1994; Williams et al.
1999) indirectly in birds of unknown status. In this
758
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assessment of
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and breeding
chronology
paper we extend this approach to the characterization of reproductive state and breeding phenology in
a population of Marbled Murrelet (Brachyramphus
marmoratus, Gmelin 1789) where individuals can only
be caught in large numbers off the nest (and thus where
these birds are all of unknown breeding status). The
Marbled Murrelet is extremely difficult to study because
it nests in relatively inaccessible habitats: large trees
in coastal old growth forest. Concern for the Marbled
Murrelet has grown in recent years due to observed
population declines over much of the species range
(near-shore waters of Alaska to central California)
associated with fragmentation and harvesting of nesting habitats through forestry development (Nelson
1997). However, a lack of basic information on population and breeding biology has hampered the development of management and conservation protocols for
this species (Ralph et al. 1995).
Hormonal and physiological changes associated
with the avian reproductive cycle have been well characterized in a wide range of both domestic and free-living
species (Wingfield & Farner 1978; Dawson 1983; Mays,
Vleck & Dawson 1991; Cockrem & Seddon 1994;
Christians & Williams 1999). This allows the identification of stage-specific hormone(s) or metabolite
profiles that might be used to characterize the reproductive state of individuals in a mixed population
(e.g. in most vertebrates, androgens and gonadotropins
are elevated during early stages of breeding, such as
courtship and oogenesis, whereas prolactin is elevated
during later periods coincident with lactation or broodrearing; Goodman 1999; Williams 1999). In this paper
we focus on the two main egg yolk precursors of
oviparous (non-mammalian) vertebrates: vitellogenin
(VTG) and very low-density lipoprotein (VLDL). Both
precursors are synthesized and secreted by the liver in
response to oestradiol (E2), and are greatly elevated
in the circulation during egg production (Griffin &
Hermier 1988; Burley & Vadehra 1989; Mitchell &
Carlisle 1991). We tested whether measurement of
plasma levels of VTG and VLDL could be used to
detect evidence of egg production in individual birds
(distinguishing egg-producing, or fecund; females from
non-egg-producing individuals), and to provide information on breeding phenology at the population level in
free-living Marbled Murrelets captured off the nest. We
validated this methodology using plasma samples from
a related species, Cassin’s Auklets (Ptychoramphus
aleuticus, Pallas 1811), where breeding chronology could
be determined (at least at the population level) by directly
monitoring large numbers of nest burrows at the colony.
Materials and methods
© 2000 British
Ecological Society,
Functional Ecology,
14, 758 –765
,   
  
Cassin’s Auklets were captured on Triangle Island
(British Columbia, Canada; 50°52′ N 129°05′ W) in
March–June 1997 and 1998. Birds were intercepted
as they left nesting burrows in the colony using soft
plastic ‘pheasant’ nets at the base of nesting slopes
(all captures between 0200 and 0530 h). Although the
reproductive status of individual Cassin’s Auklets was
unknown, timing of egg-laying, incubation and chickrearing in the colony was monitored directly by
researchers checking large numbers (~400) of active
burrows, allowing accurate breeding chronologies to
be determined (Triangle Island Research Station,
unpublished data). Marbled Murrelets were captured
at Desolation Sound, British Columbia (50°05′ N,
124°40′ W) between May–August 1996 and 1997. In
1996, a floating mist-net system (Kaiser et al. 1995)
was used to capture adults as they flew between inshore
marine foraging areas and forest breeding areas,
through Theodosia Inlet, Desolation Sound around
dawn (0400–0700 h) and dusk (2000 – 2300 h). In 1997,
in addition to mist-netting, a ‘night-lighting’ technique
(Whitworth et al. 1997) was used to capture birds on
the water at night (2300 –0500 h) in Desolation Sound.
Both Marbled Murrelets and Cassin’s Auklets were
blood sampled and sexed following the methods
described in Vanderkist et al. (1999), and body mass
was recorded for all captured individuals.
    ’ 

Descriptive statistics were used to construct an interval
(mean ± 2 SD) around mean laying and hatching dates
(Triangle Island Research Station, unpublished data);
laying, mean 9 April (range 18 March to 30 April);
hatching, mean 12 May (range 24 April to 30 May).
The intervals for laying and hatching periods overlapped by about 7 days, so the mid-point of the overlap (27 April) was used as the boundary between the
stages of laying and incubation. For the chick-rearing
stage, an average fledging period of 41–50 days was
used (Manuwal & Thoresen 1993). In 1998, plasma
samples of presumed prelaying Cassin’s Auklets
were also obtained, collected as early in the season as
possible (24–28 March) before any eggs were detected
in nest burrows.
   
 -  - 
Plasma levels of VTG and VLDL were assayed with
diagnostic kits (Zn, Cat no. 435–14909; Triglyceride
E Kit, Code no. 432–40201, Wako Pure Chemical
Industries Ltd, Richmond, VA, USA) following the
methods of (Mitchell & Carlisle 1991). Vitellogenic
zinc was used as an index for VTG, and total triglyceride as a measure of VLDL (see also Williams &
Christians 1997; Williams & Martyniuk 2000). Interassay coefficients of variation for VLDL and VTG were
4·3% and 8·6%, respectively, and intra-assay coefficients
FEC475.fm Page 760 Thursday, November 16, 2000 8:51 AM
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B. A. Vanderkist
et al.
of variation were 3·1% and 5·9%, respectively. In some
cases, both VTG and VLDL could not be measured
due to small plasma sample volume.
Plasma levels of VTG and VLDL in male birds
were used to assign an upper limit for classification of
‘non-egg-producing’ individuals, since males normally
do not produce VTG and they have basal VLDL levels
(Mitchell & Carlisle 1991; Williams & Christians 1997).
The highest values of VTG and VLDL observed in male
Cassin’s Auklets were 0·11 µg ml–1 Zn and 1·36 m
triglyceride, respectively. For male Marbled Murrelets,
the highest values of VTG and VLDL were 0·26 µg ml–1
Zn and 6·62 m triglyceride, respectively. To be
conservative, the maximum yolk precursor values
obtained for males were doubled and used as limits
to classify females as either egg-producing or nonegg-producing birds.
 
All statistical analyses were performed with SAS
(version 6·11) or Minitab (version 11) statistical software. Both parametric and non-parametric statistical
analyses were used depending upon the results of
normality tests for the data being examined. Values
are presented as the mean ± standard deviation unless
otherwise stated.
Results
 ’  
Cassin’s Auklets classified as egg-producing birds
were detected from 24 March to 30 April, and capture dates for these birds closely matched the range of
laying dates obtained by direct observation of nest
burrows (18 March to 30 April). Plasma VTG levels
were on average two- and three-fold higher, respectively,
during the defined egg-laying period compared to the
prelaying and incubation periods (Fig. 1). The proportion of females classified as egg-producers using
VTG differed significantly between breeding stages
(Fisher’s exact test, χ22 = 11·5, P < 0·01), with the
highest proportion recorded during the laying period
(Table 1).
Fig. 1. Variation in plasma vitellogenin levels by defined
reproductive stage in Cassin’s auklets. Sample sizes for each
stage are given at the top of each box plot.
The proportion of females classified as egg-producers
using VLDL also differed significantly between reproductive stages (Fisher’s exact test; χ 22 = 6·9 P < 0·05;
Table 1). The absolute number of female Cassin’s
Auklets classified as egg-producing was less using
VLDL than using VTG (Table 1). However, the proportion of females classified as egg-producing were
only marginally significantly different between the
two methods (χ 21 = 3·85, P = 0·05).
 
No Marbled Murrelet females were classified as eggproducing birds using either VTG or VLDL in 1996.
Birds were only captured by mist-netting in 1996 and,
more importantly, the range of sample dates was much
later in 1996 (June – August) than in 1997 (May–August;
Fig. 2). In contrast, in 1997 there was much greater
interindividual variation in plasma VTG levels in
female Murrelets (Table 2) when the night-lighting
technique permitted the capture of birds earlier in the
season. In 1997, Marbled Murrelet females classified
as egg-producing birds were detected between 14 May
and 3 July based on plasma VTG levels, clearly indicating the putative egg-laying period for this species.
Only 2 out of a total of 23 females classified as eggproducing using VTG were captured by mist-netting.
As with Cassin’s Auklets, use of plasma VTG
detected a greater absolute number of egg-producing
Table 1. Levels of VTG-Zn and VLDL and the proportion of egg-producing female Cassin’s Auklets within each
reproductive stage. Limits for classifying individuals as egg-producing using VTG-Zn and triglyceride were 0·22 µg ml–1 Zn
and 2·7 m triglyceride, respectively
Yolk precursor
VTG (µg ml–1 Zn)
© 2000 British
Ecological Society,
Functional Ecology,
14, 758 –765
VLDL (m TTRIG)
Reproductive
stage
n
Mean ± SD (range)
% elevated VTG-Zn
n
Mean ± SD (range)
% elevated VLDL
Prelaying
Laying
Incubation
Chick-rearing
10
22
19
8
0·73 ± 1·26 (0·00 – 4·07)
1·66 ± 1·97 (0·00 – 6·56)
0·55 ± 1·37 (0·00 – 5·70)
0·10 ± 0·15 (0·00 – 0·43)
30
55
16
0
9
24
18
8
1·54 ± 0·73 (0·75 –2·74)
7·40 ± 11·53 (0·54–42·21)
3·15 ± 7·91 (0·59 –34·64)
0·98 ± 0·36 (0·423–1·453)
11
33
11
0
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Physiological
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and breeding
chronology
Fig. 2. Variation in plasma vitellogenin levels in relation to
capture date in Marbled Murrelets, in (a) 1996 (n = 24) and
(b) 1997 (n = 107).
females than did VLDL, but there was no significant
difference in the proportion of egg-producing females
detected using either yolk precursor (χ21 = 2·44, P > 0·1).
   
   
Plasma levels of VTG and VLDL were positively correlated in female Cassin’s Auklets (F1,53 = 118·3,
P < 0·001, r2 = 0·69), in female Marbled Murrelets
captured in 1997 (F1,101 = 256·42, P < 0·001, r 2 = 0·72),
but not in female Marbled Murrelets captured in
1996 (F1,22 = 1·59, P > 0·2, r 2 = 0·067; Fig. 3). Some
individuals of both species would have been classified as egg-producers using VTG as an index, but not
if using VLDL (Fig. 3). Three patterns of VTG–VLDL
levels were defined: (a) elevated VTG and VLDL;
(b) elevated VTG only; and (c) neither VTG or VLDL
elevated (no females from either species displayed
Fig. 3. Relationship between plasma vitellogenin and
plasma VLDL in individual birds, for (a) Cassin’s Auklets,
(b) Marbled Murrelets in 1996 and (c) Marbled Murrelets
in 1997. Dashed lines are the egg-producing limits for each
yolk precursor as described in the text.
elevated levels of VLDL only). Two female Marbled
Murrelets with high VTG but not VLDL (pattern b)
were known to be carrying eggs at their time of capture,
since an egg was physically detected in the oviduct
during banding.
There was a significant difference in body mass
between individuals with the defined patterns of eggprecursor levels for female Cassin’s Auklets (F2,44 = 9·63,
P < 0·001; Fig. 4). Females with elevated levels of
both VTG and VLDL (pattern a) and females with
elevated levels of VTG only (pattern b) were significantly heavier than females with basal levels of both
VTG and VLDL (pattern c; Bonferonni-adjusted
comparison, df = 44, tcrit = 2·49). The 95% confidence
intervals for the body mass of egg-producing and
Table 2. Levels of VTG-Zn and VLDL for 1997 Marbled Murrelets by 30-day intervals and proportions of egg-producing females within each capture
method using either VTG-Zn or VLDL as an index
Yolk precursor
VTG-Zn (µg ml–1)
VLDL (mM TTRIG)
% elevated VTG-Zn
14 May to 12 June
©
132000
June British
to 12 July
Ecological
Society,
13
July to 11
August
Functional
Ecology,
Total
14, 758 –765
% elevated VTG-Zn
Mean ± SD (range)
Night-lighting
Mist-netting
Mean ± SD (range)
Night-lighting
Mist-netting
2·58 ± 2·7 (0·00 – 8·67)
0·83 ± 1·94 (0·00 – 8·13)
0·11 ± 0·11 (0·00 – 0·38)
1·08 ± 2·10 (0·00 – 8·67)
60 (15 / 25)
100 (6 / 6)
0 (0 / 24)
38 (21 / 55)
0 (0 / 2)
4 (2 / 45)
0 (0 / 5)
4·0 (2 / 52)
17·3 ± 24·5 (0·7 – 80·4)
5·2 ± 14·4 (0·5 – 81·2)
1·1 ± 0·4 (0·5 – 2·9)
6·7 ± 16·3 (0·5 – 81·2)
38 (10/26)
57 (4/7)
0 (0/32)
22 (14/65)
0 (0/2)
0 (0/44)
0 (0/6)
0 (0/52)
FEC475.fm Page 762 Thursday, November 16, 2000 8:51 AM
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B. A. Vanderkist
et al.
Fig. 4. Variation in body mass for (a) female Cassin’s Auklets and (b) Marbled
Murrelets in relation to the patterns of yolk precursor levels: (A) both VTG-Zn and
VLDL levels elevated, (B) only VTG-Zn levels elevated and (C) neither VTG-Zn or
VLDL levels elevated. Numbers at top of graphs are sample sizes, error bars are
standard deviations.
non-egg-producing female Cassin’s Auklets were 179 –
190 g, and 166 –174 g, respectively. Similarly, female
Marbled Murrelets captured in 1997 showed a significant difference in body mass between patterns of yolkprecursor levels (F2,100 = 35·30, P < 0·001; Fig. 4). Again,
females with elevated levels of VTG and VLDL
(pattern a) and females with elevated levels of only
VTG (pattern b) were significantly heavier than those
with low levels of both VTG and VLDL (pattern c;
df = 101, tcrit = 2·43). The 95% confidence intervals for
the body mass of egg-producing, non-egg-producing
and male Marbled Murrelets were 225–245 g, 195–
202 g and 198 – 202 g, respectively.
Discussion
© 2000 British
Ecological Society,
Functional Ecology,
14, 758 –765
In this study we tested the validity of using indirect,
physiological, measures to provide information on
reproductive state and breeding phenology, in a situation where these data could not be obtained through
more traditional approaches (direct observation of
nests). We focused on plasma levels of the two main
yolk precursors, vitellogenin and very low-density
lipoprotein because we predicted that these would
provide highly specific indicators of a single repro-
ductive stage: elevated levels should occur in eggproducing females, but not in non-breeders, males or
females at other breeding stages. Our data for Cassin’s
Auklets, where breeding could be directly monitored,
support this assumption and confirm the validity of
this technique. In this species, the highest levels of
VTG and VLDL were observed during the defined
laying period (constructed from direct burrow observations), and the highest proportion of birds were
identified as ‘egg-producing’ during this period (see
Fig. 1). In Marbled Murrelets, where the status of
individuals and timing of breeding were completely
unknown, measurement of plasma VTG and VLDL
clearly identified the putative egg-laying period (from
mid-May to early July, see Fig. 2). These data also
indicate that Marbled Murrelets have a single, protracted breeding season, with no evidence of a bimodal
distribution which would be associated with multiple
broods (second clutches). In addition, since we obtained
data on VTG and VLDL for individual birds, this technique can accurately assign reproductive status to some
individuals (egg-laying females). This may be valuable
where studies need to target fecund females (e.g. for
radio-transmitter attachment in behavioural studies).
Both yolk precursors, VTG and VLDL, were
measured to assess whether these were equally reliable
as indices of egg production. In fact, VTG detected
greater absolute numbers of egg-producing females
than did VLDL, in both Cassin’s Auklets and Marbled
Murrelets. Some evidence was therefore obtained that
VTG and VLDL were elevated in plasma for different
lengths of time in relation to egg formation, because
there was not 100% agreement when classifying a
female as an egg-producing bird. For example, the
two Marbled Murrelets carrying eggs at their time of
capture had elevated levels of VTG, but not VLDL;
these birds would have been misclassified as ‘nonegg-producing’ females had we only considered
VLDL. This result is somewhat surprising given that
both yolk precursors are essential for yolk formation. In birds, triglyceride-rich lipoproteins (VLDL)
are a significant component of egg-yolk (66% of hen’s
egg yolk, Griffin & Hermier 1988; Williams 1999).
Furthermore, both yolk-precursors are under the
same oestrogenic control (Wiskocil et al. 1980; Wahli
et al. 1981; Burley & Vadehra 1989; Mitchell & Carlisle
1991), and are also believed to share the same oocyte
receptor in birds (Stifani et al. 1990; Elkin et al. 1995).
However, very little is known about yolk precursor
dynamics during the rapid yolk development period,
or about the relative half-life of each precursor, and
this may explain the discrepancies found. An alternative explanation is that VLDL provides a ‘noisier’
signal for egg-production than does VTG. In contrast
to VTG, non-yolk VLDL does occur in non-egg-laying
birds, i.e. the baseline value for non-breeders is not
zero (as it is for VTG). Plasma levels of this non-yolk
VLDL can vary with diet and physiological state (e.g.
fasting vs fattening). However, diurnal variation in
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Ecological Society,
Functional Ecology,
14, 758 –765
plasma triglycerides or variation due to feeding/fasting
is typically in the range 0·5–3·0 mg ml–1 (e.g. JenniEierman & Jenni 1996, 1997). This is similar to the
range of variation we report for birds during chickrearing (see, for example, Fig. 3) when no egg formation was occurring but when there was most likely
natural variability in whether birds had fed recently
or not. Variation in VLDL levels in non-breeders
therefore occurs at an order of magnitude lower than
VLDL levels in laying birds (10–40 mg ml–1). Thus,
although there were almost certainly differences in
the physiological state of the birds we caught (fed vs
fasted), as well as between Cassin’s Auklets and Marbled
Murrelets, we doubt that this confounds interpretation of VLDL in relation to egg-laying to any great
extent. Plasma VTG levels are independent of diet
(T. D. Williams unpublished data for Zebra Finches),
and do not appear to show marked diurnal variation
(Redshaw & Follett 1976; T. D. Williams, unpublished
data), reducing the potential confounding effects of
time of capture and physiological state (fed vs fasted)
for this precursor. Our data, and those for other studies,
therefore suggest that VTG provides the more accurate
and reliable index of egg production. Nevertheless,
there are two advantages of using VLDL: (a) VLDL
is easier to measure than VTG, and (b) the VLDL assay
requires smaller plasma sample volumes (so this yolk
precursor may have value in certain studies).
Few previous studies have analysed plasma levels
of VTG and VLDL in relation to egg formation in
free-living or domesticated birds. In chickens (Mitchell
& Carlisle 1991) and captive-breeding Zebra Finches
(Taeniopygia guttata; Williams & Christians 1997)
plasma yolk precursors are elevated during laying, but
are undetectable in immatures, non-breeding females
and males. Similarly, Berry, Millar & Louw (1979)
reported elevated levels of phosvitin (a precursor of
VTG) and triglyceride in breeding but not non-breeding
female Cape Cormorants (Phalacrocorax capensis).
In free-living European Starlings (Sturnus vulgaris),
plasma VTG was significantly higher in laying birds
compared with prebreeders (sampled 1–4 days before
egg-laying), and had decreased to less than 10% of
peak values only 2 days after clutch completion
(Christians & Williams 1999). These studies confirm
that elevated plasma VTG levels are highly specific
to egg formation and laying. Marbled Murrelets and
Cassin’s Auklets lay a single egg clutch, and rapid yolk
development, during which yolk precursors should
be elevated, takes about 14 days in Cassin’s Auklets
(Astheimer & Grau 1990; no data available for
Murrelets).
In our study, body mass varied significantly with
plasma VTG levels, confirming our characterization
of ‘egg-producing’ females. For Marbled Murrelets,
the difference in mean body mass between birds with
low levels of VTG (pattern C) and those with high
levels of VTG (patterns A and B) was in the range of
40 – 46 g, consistent with the expected egg mass of
this species (36–41 g, Nelson 1997). Body mass itself
therefore proved to be a somewhat useful indicator
of egg-production in Marbled Murrelets, with eggproducing females being significantly heavier than both
males and non-egg-producing females. This finding
might be useful for researchers who wish to maximize
the probability of marking, sampling or manipulating
females within a capture sample, e.g. for the attachment of radio-transmitters on breeding individuals
which could then be tracked back to a nest-site.
In one year of our study (1996), when Marbled
Murrelets were captured late in the season, from June
to August, using floating mist-nets, we detected no
egg-producing females. Night-lighting allowed us to
capture birds much earlier in the season (in 1997),
and consequently many more birds were identified as
‘egg-producing’. Of 77 female Marbled Murrelets
captured by mist-netting at Theodosia Inlet, Desolation Sound during the 2 years of this study, only two
were classified as egg-producing using VTG. This is
unlikely to represent a bias due to different capture
techniques, rather it simply is due to the fact that
mist-netting could not be employed as early in the
season as night-lighting (when egg-production would
be more probable). In 1997, egg-producing Marbled
Murrelets were detected from 14 May to 3 July. These
dates agree reasonably well with Marbled Murrelet
chronology estimates in British Columbia (Hamer &
Nelson 1995), that predicted incubation to last from
2 May to 4 July, assuming a 14-day rapid yolk development period in Marbled Murrelets as in Cassin’s
Auklets (Astheimer 1986). However, because our
night-lighting sampling effort from 23 June to 12 July
was greatly reduced (associated with a switch to mistnetting), it is possible that during this time there were
still significant numbers of individuals producing eggs
that would have been detected had we maintained
our night-lighting effort. Similarly, juvenile Marbled
Murrelets were first observed at sea on 27 June in 1997
(C. Lougheed, personal communication). Assuming
58 days between laying and fledging (Hamer & Nelson
1995), this indicates that some birds started laying
before our first captures. Further sampling, with earlier
capture of birds, will hopefully refine our estimates of
breeding chronology of Marbled Murrelets using this
technique, as well as addressing the issue of interannual variation in breeding phenology.
In summary, measurement of plasma VTG and
VLDL provided useful indicators of reproductive state
(egg production), although our data indicate that
VTG provides the more robust, reliable index. Use of
yolk precursors for this type of analysis has the additional advantage that these involve relatively simple
diagnostic assays (cf. more involved ELISA (enzymelinked inmmunosorbent assay) or RIA (radioimmunoassay) techniques). A limitation of this general
approach is that single measures of hormone or metabolite profiles only provide a static picture of dynamic
physiological systems. We could only assess whether
FEC475.fm Page 764 Thursday, November 16, 2000 8:51 AM
764
B. A. Vanderkist
et al.
females were or were not producing an egg at the
time of capture. Owing to interindividual variation in
the timing of breeding among our samples of birds,
there were probably females that had already laid
eggs and were well into later stages of reproduction,
along with females that may have been preparing
to initiate egg formation. Both of these groups of
females would be classified as non-egg-producing
birds, but were not necessarily non-breeding adults.
Circulating levels of yolk precursors therefore provide a minimum estimate of the proportion of breeding females in a given capture sample. Nevertheless,
in our study, indirect, physiological assessment of
reproductive state provided valuable information on
the breeding biology of Marbled Murrelets which
would have been difficult to obtain in any other way
(e.g. breeding phenology, single vs multiple-clutch
breeding pattern, as well as reproductive status for
some individuals at the time of capture). Combining measurement of yolk precursors with that of
other hormones (e.g. prolactin) might resolve some
of the problems of underestimating the number of
fecund females. Individuals sampled during the defined
egg-laying period with low VTG but high prolactin
(indicating ongoing parental care) are likely to have
previously laid an egg (i.e. to have completed egg
formation), whereas individuals with a low VTG:
low prolactin profile are more likely to be true nonbreeders. Finally, given the widespread occurrence and
sex-specific nature of vitellogenin this approach should
be applicable to all oviparous vertebrates (see, e.g.,
Takemura & Oka 1998; Heppell & Sullivan 2000).
Acknowledgements
We thank the various funding agencies that made this
work possible: Forest Renewal BC through Science
Council BC, Ministry of Forests, BC, Natural Sciences
and Engineering Research Council of Canada,
Canadian Wildlife Service, MacMillan-Bloedel Ltd,
TimberWest Forest Ltd, International Forest Products
Ltd, Western Forest Products Ltd and Pacific Forest
Products Ltd. We also thank Laura Jones and Kerry
Woo for their tireless efforts in monitoring nest burrows
and collecting blood samples at Triangle Island, and
thank Connie Smith for providing data on Cassin’s
Auklet breeding biology collected there. We acknowledge the many hard-working members of the Marbled
Murrelet research project, particularly Cecilia Lougheed,
Gary Kaiser and Dr Fred Cooke. Special thanks to
Julian Christians and Dr Cindy Hull for their comments and suggestions for this manuscript.
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Received 9 September 1999; revised 12 April 2000; accepted
11 May 2000